Project Abstract A reversible non-hormonal male contraceptive that is reliable, safe, and easy to use will provide a significant option for men who want to take an active role in family planning. The current primary choices for men are vasectomy and condoms, but each has problems with reversibility and surgical risk, or compliance, respective- ly. There is currently a paucity of druggable targets in drug development for reversible non-hormonal male con- traception. Protein kinases that are critical for cell cycle progression during spermatogenesis that are drugga- ble offer promise as targets for contraceptive development. In this regard, cyclin-dependent protein kinase 2 (CDK2) is such a target. CDK2-/- knockout mice are viable but sterile. CDK2 through its interaction with cyclins A and A1 targets retinoblastoma protein (pRB) and p53 for phosphorylation. Both of these proteins are present in human and murine testis, and play critical roles in testis development, and in regulation of transition through the spermatogenic cell cycle. Therefore, a drug that effectively inhibits CDK2 should block spermatogenesis, thereby causing infertility due to lack of sperm. Since CDK2 is an enzyme that effects meiotic division, once the drug is no longer taken, meiosis should resume and sperm production recovers. The primary hypothesis for this research is that novel small molecule specific inhibitors of CDK2 can be developed as novel non-hormonal reversible male contraceptive agents. To prove this hypothe- sis, the following specific aims will be achieved: Specific Aim 1. Identify and rank known inhibitors of CDK2 that reversibly inhibit spermatogenesis in rats. Specific Aim 2. Develop novel CDK2 allosteric and type I and II kinase inhibitors as leads to improve CDK2 specificity, in vivo anti-spermatogenic efficacy, and minimize side effects. Specific Aim 3. Demonstrate proof-of-concept reversible contraceptive efficacy of CDK2 inhibitors To achieve specific aims, we will undertake two lines of research to develop novel CDK2 inhibitors as male contraceptives. First, we will improve the CDK2 specificity for potent nanomolar type I and type II kinase inhibitors that we recently developed. This approach of building in specificity has worked for us before. Second, we will pursue the development of novel CDK2 modulators via an allosteric site that we recently discovered, as these may provide greater opportunities to have CDK2-specific selectivity and minimize cross reactivity with other members of the kinase family. In order to be a successful and viable male contraceptive agent, efficacy, safety, and recovery of fertility will need to be comparable to the oral female contraceptive pill. Based on our experience and success in ongoing contraceptive development projects, one of which is currently in pre-clinical development under FDA guidance, we expect this project to produce novel CDK2 inhibitors with high promise to meet the goal of a safe, effective, and easy to use reversible male contraceptive agent.